Neoplastic cells carry upon their surfaces antigens which distinguish them from their normal counterparts, with chemically induced neoplasms frequently expressing unique antigens, denoted tumor-specific transplantation antigens (TSTA). The present research has demonstrated that TSTA activity can be extracted from intact tumor cells using single-phase solutions of n-butanol. The extraction technique is not cytotoxic, as judged by the ability of extracted cells to proliferate in vitro and in vivo. Crude butanol extracts are immunogenical in immunoprotection assays and evoke a positive delayed hypersensitivity (DH) response in immune mice. However, crude butanol extracts do not contain measurable amounts of alloantigenic activity. It has also been shown that low concentrations of octylglucoside also removes immunogenic TSTA activity from intact cells without affecting viability. Thus, two noncytolytic extraction techniques have been developed for the preparation of TSTA moieties which are stable in aqueous media. Present research involves isolation and purification of the TSTA molecules from a panel of 3-MCA-induced fibrosarcomas of C3H/HeJ mice. The butanol-extracted materials will be fractionated by isoelectric focusing, molecular sieve chromatography, polyacrylamide gel electrophoresis, hydroxyapatite chromatography and affinity chromatography. The presence of an active TSTA moiety will be assessed by the induction of immunoprotection and by evocation of a DH response. During purification, the TSTA-active principles will be characterized with respect to charge, apparent size, association with lipids and degree of glycosylation. Recent progress has demonstrated the TSTAs from the tumors MCA-F, MCA-D and MCA-2A have very similar physicochemical properties, including an isoelectric pH of 6.4\-\6.6 and a native molecular weight of 150,000 daltons (150 kd). The 150 kd antigen is free of alloantigenic activity, but does bear epitopes which react in ELISA with monoclonal reagents specific for the qp70 and p15(E) antigens from MuLV. Dissociation of the 150 kd antigens at pH 3 yield two major fractions, one a 70 kd which is not immunoprotective and another at 35 kd that bears the tumor specific epitope.